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HPLC System (hplc + system)
Selected AbstractsA simple and rapid high-performance liquid chromatographic (HPLC) method for 5-fluorouracil (5-FU) assay in plasma and possible detection of patients with impaired dihydropyrimidine dehydrogenase (DPD) activityJOURNAL OF CLINICAL PHARMACY & THERAPEUTICS, Issue 4 2004J. Ciccolini PharmD PhD Summary Background:, Dihydropyrimidine dehydrogenase (DPD) gene polymorphism may lead to severe toxicity with 5-fluorouracil (5-FU), a major anticancer drug extensively used in clinical oncology. Drug monitoring combined with early detection of patients at risk would enable timely dose adaptation so as to maintain drug concentrations within a therapeutic window. However, the best method to identify such patients remains to be determined. Objective:, The aim of this study was to develop a rapid and simple high-performance liquid chromatographic (HPLC) method for estimating uracil/dihydrouracil (U/UH2) ratio in plasma, as an index of DPD status, and for assaying 5-FU as part of drug level monitoring. Method:, Assay of 5-FU, and U/UH2 detection were performed on a HPLC system equipped with UV detector. Analytes were separated at room temperature using a 5 ,m particles, 25 cm RP-18 X-Terra column. The mobile-phase consisted of a KH2PO4 salt solution (0·05 m) + 0·1% triethylamine (TEA) pumped at 0·4 mL/min. Detection of 5-FU and 5-bromouracil were performed at 254 nm; U and UH2 elution was monitored at 210 nm. Results:, The method was sensitive and specific for assaying 5-FU within the 5,500 ng/mL concentration range, which covers exposure levels currently met in clinical practice. The method was simple, and relatively cheap, and rapid, with an analytical run time of about 30 min. Data from a patient with 5-FU toxicity suggest that the method was capable of identifying DPD metabolic phenotype in cancer patients, based on measurement of plasma U/UH2 ratio. Conclusion:, The method described should be suitable both for detecting patients at high risk of 5-FU toxicity, and for drug level monitoring during chemotherapy. [source] PURIFICATION AND CHARACTERIZATION OF BACTERIOCIN FROM WEISSELLA PARAMESENTEROIDES DFR-8, AN ISOLATE FROM CUCUMBER (CUCUMIS SATIVUS)JOURNAL OF FOOD BIOCHEMISTRY, Issue 5 2010AJAY PAL ABSTRACT Bacteriocin from Weissella paramesenteroides DFR-8 isolated from cucumber (Cucumis sativus) was purified by using only two steps, viz., pH-mediated cell adsorption,desorption method and gel permeation chromatography. A single peak observed in the purity check by analytical Reverse Phase-High Performance Liquid Chromatography (Waters 600 analytical HPLC system, Milford, MA) and a single band (molecular weight,3.74 kDa) shown on SDS-PAGE analysis strongly indicated the homogeneity of the bacteriocin preparation. Treatment with proteolytic enzymes abolished the antimicrobial activity indicating the proteinaceous nature of bacteriocin. The purified bacteriocin exhibited a broad inhibitory spectrum against foodborne pathogens and spoilage microorganisms, including gram-negative bacteria such as Salmonella typhimurium, Vibrio parahaemolyticus, Aeromonas hydrophila and Listeria monocytogenes. Response surface methodology was employed to study the interactive effect of temperature and pH on bacteriocin activity, and a regression equation was developed. The bacteriocin retained full activity after storage at,20C for 90 days, while partial and complete activity loss was observed when stored at 4 and 37C, respectively. PRACTICAL APPLICATION In recent years, bacteriocins of lactic acid bacteria have gained much attention as food biopreservatives because of their origin from generally regarded as safe organisms. In spite of various bacteriocins studied worldwide, studies on bacteriocins of Weissella paramesenteroides remain rare. The present work involves the purification of bacteriocin up to absolute homogeneity from W. paramesenteroides, an isolate first time reported from cucumber (Cucumis sativus). The purified bacteriocin (molecular weight ,3.74 kDa) was found to inhibit a large number of foodborne pathogens, including Listeria monocytogenes, which is resistant to commercially available bacteriocin, i.e., nisin. The application of central composite rotatable design enabled us to design a regression equation from which the residual activity of bacteriocin can be predicted at any given conditions of temperature and pH within the experimental domain. The broad inhibitory spectrum and thermostability of bacteriocin suggest its potential application in food preservation. [source] HPLC methods for the purification of [11C]-labelled radiopharmaceuticals: reversal of the retention order of products and precursorsJOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS, Issue 5 2009Szabolcs Lehel Abstract Preparative HPLC methods have been developed for a number of [11C]-methylated PET tracers, which enable elution of the labelled compounds prior to their precursors, thus reducing the overall synthesis time and avoiding contamination of the final product with precusor. This reversal of retention order has been achieved for [11C]DASB, [11C]raclopride, [11C]FLB 457, [11C]carfentanil, and 2-fluoro-[N -methyl- 11C]apomorphine, enabling collection of the purified radiopharmaceuticals from the HPLC system after 5,7,min. Furthermore, by using ethanol as the organic modifier, residual solvent analysis prior to human injection could be avoided and three of the radiopharmaceuticals could be injected directly following simple dilution and sterile filtration. Copyright © 2009 John Wiley & Sons, Ltd. [source] Quantitative analysis of EO9 (apaziquone) and its metabolite EO5a in human plasma by high-performance liquid chromatography under basic conditions coupled to electrospray tandem mass spectrometryJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 10 2006Liia D. Vainchtein A sensitive and specific LC-MS/MS assay for the quantitative determination of EO9 and its metabolite EO5a is presented. A 200-µl human plasma aliquot was spiked with a mixture of deuterated internal standards EO9- d3 and EO5a- d4 and extracted with 1.25 ml ethyl acetate. Dried extracts were reconstituted in 0.1 M ammonium acetate,methanol (7 : 3, v/v) and 25 µl-volumes were injected into the HPLC system. Separation was achieved on a 150 × 2.1 mm C18 column using an alkaline eluent (1 mM ammonium hydroxide,methanol (gradient system)). Detection was performed by positive ion electrospray followed by tandem mass spectrometry. The assay quantifies a range from 5 to 2500 ng/ml for EO9 and from 10 to 2500 ng/ml EO5a using 200 µl of human plasma samples. Validation results demonstrate that EO9 and EO5a concentrations can be accurately and precisely quantified in human plasma. This assay will be used to support clinical pharmacologic studies with EO9. Copyright © 2006 John Wiley & Sons, Ltd. [source] Identification of Kaempferol as a Monoamine Oxidase Inhibitor and Potential Neuroprotectant in Extracts of Ginkgo Biloba LeavesJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 4 2000B. D. SLOLEY The effects of Ginkgo biloba leaf extract on rat brain or livermonoamine oxidase (MAO)-A and -B activity, biogenic amine concentration in nervous tissue, N -methyl- d -aspartate (NMDA)- and N -(2-chloroethyl)- N -ethyl-2-bromobenzylamine (DSP-4)-induced neurotoxicity and antioxidant activity was investigated to determine the effects of the extract on monoamine catabolism and neuroprotection. Ginkgo biloba leaf extract was shown to produce in-vitro inhibition of rat brain MAO-A and -B. The Ginkgo biloba extract was chromatographed on a reverse-phase HPLC system and two of the components isolated were shown to be MAO inhibitors (MAOIs). These MAOIs were identified by high-resolution mass spectrometry as kaempferol and isorhamnetin. Pure kaempferol and a number of related flavonoids were examined as MAOIs in-vitro. Kaempferol, apigenin and chrysin proved to be potent MAOIs, but produced more pronounced inhibition of MAO-A than MAO-B. IC50 (50% inhibition concentration) values for the ability of these three flavones to inhibit MAO-A were 7 times 10,7, 1 times 10,6 and 2 times 10,6m, respectively. Ginkgo biloba leaf extract and kaempferol were found to have no effect ex-vivo on rat or mouse brain MAO or on concentrations of dopamine, noradrenaline, 5-hydroxytryptamine and 5-hydroxyindoleacetic acid. Kaempferol was shown to protect against NMDA-induced neuronal toxicity in-vitro in rat cortical cultures, but did not prevent DSP-4-induced noradrenergic neurotoxicity in an in-vivo model. Both Ginkgo biloba extract and kaempferol were demonstrated to be antioxidants in a lipid-peroxidation assay. This data indicates that the MAO-inhibiting activity of Ginkgo biloba extract is primarily due to the presence of kaempferol. Ginkgo biloba extract has properties indicative of potential neuroprotective ability. [source] Liquid,liquid,liquid microextraction followed by HPLC with UV detection for quantitation of ephedrine in urineJOURNAL OF SEPARATION SCIENCE, JSS, Issue 18 2008Habib Bagheri Abstract Liquid,liquid,liquid microextraction (LLLME) in combination with HPLC and UV detection has been used as a sensitive method for the determination of ephedrine in urine samples. Extraction process was performed in a homemade total glass vial without using a Teflon ring, usually employed. Ephedrine was first extracted from 3.5 mL of urine sample (pH 12) into a microfilm of toluene/benzene (50:50). The analyte was subsequently back extracted into an acidic microdrop solution (pH 2) suspended in the organic phase. The extract was then injected into the HPLC system directly. An enrichment factor of 137 along with a good sample clean-up was obtained under the optimized conditions. The calibration curve showed linearity in the range of 0.01,50 mg/L with regression coefficient corresponding to 0.998. The LODs and LOQs, based on a S/N of 3 and 10, were 5 and 10 ,g/L, respectively. The method was eventually applied for the determination of ephedrine in urine sample after oral administration of 5 mg single dose of drug. [source] Separation of triacylglycerols in a complex lipidic matrix by using comprehensive two-dimensional liquid chromatography coupled with atmospheric pressure chemical ionization mass spectrometric detectionJOURNAL OF SEPARATION SCIENCE, JSS, Issue 8 2006Paola Dugo Abstract The present investigation describes the employment of a comprehensive 2-D HPLC system, based on the combination of a silver ion and an RP column, for the characterization of the triacylglycerol (TAG) fraction of a very complex lipidic sample: donkey milk fat. The TAGs were grouped on the resulting bidimensional contour plot according to their double bond numbers (aligned along vertical bands) and according to their partition numbers (aligned along horizontal bands). Peak assignment was supported by using atmospheric pressure chemical ionization mass spectrometric (APCI-MS) detection. The combination of the enhanced resolving power of comprehensive multidimensional LC, the formation of ordered 2-D patterns, and APCI-MS detection proved to be an effective tool for the characterization of the complex matrix, enabling the separation and identification of nearly 60 TAGs. [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] Development and validation of a sensitive assay for the quantification of imatinib using LC/LC-MS/MS in human whole blood and cell cultureBIOMEDICAL CHROMATOGRAPHY, Issue 12 2009Jelena Klawitter Abstract We developed and validated a semi-automated LC/LC-MS/MS assay for the quantification of imatinib in human whole blood and leukemia cells. After protein precipitation, samples were injected into the HPLC system and trapped onto the enrichment column (flow 5 mL/min); extracts were back-flushed onto the analytical column. Ion transitions [M + H]+ of imatinib (m/z = 494.3 , 394.3) and its internal standard trazodone (372.5 , 176.3) were monitored. The range of reliable response was 0.03,75 ng/mL. The inter-day precisions were: 8.4% (0.03 ng/mL), 7.2% (0.1 ng/mL), 6.5% (1 ng/mL), 8.2% (10 ng/mL) and 4.3% (75 ng/mL) with no interference from ion suppression. Autosampler stability was 24 hs and samples were stable over three freeze,thaw cycles. This semi-automated method is simple with only one manual step, uses a commercially available internal standard, and has proven to be robust in larger studies. Copyright © 2009 John Wiley & Sons, Ltd. [source] Transport behavior of ellagic acid of pomegranate leaf tannins and its correlation with total cholesterol alteration in HepG2 cellsBIOMEDICAL CHROMATOGRAPHY, Issue 5 2009Jiaqi Lan Abstract The aim of this study was to investigate whether ellagic acid in pomegranate leaf tannins could be transported into HepG2 cells and its transport behavior. High-performance liquid chromatography coupled with a 996 photodiode array detector at 254 nm was applied. The mobile phase was an acetonitrile,water solution (containing 0.1% triethylamine, pH 3.0; 16:64, v/v, for determining ellagic acid in cells). The flow rate was 0.8 mL/min. Cells were incubated with pomegranate leaf tannins with 100 and 50 µg/mL (containing 1.71 and 0.85 µg/mL of ellagic acid, respectively) for a specific time, then lysed and sonicated in methanol to extract intracellular ellagic acid. A 10 µL aliquot of sample was injected into the HPLC system to determine ellagic acid concentration. The results showed that ellagic acid in pomegranate leaf tannins could be transported into the cells, which was in correlation with total cholesterol alteration in the cells. This is the first time that the transport behavior of ellagic acid through HepG2 cells in vitro has been comprehensively demonstrated. Copyright © 2008 John Wiley & Sons, Ltd. [source] Chiral stationary phase covalently bound with a chiral pseudo-18-crown-6 ether for enantiomer separation of amino compounds using a normal mobile phaseCHIRALITY, Issue 3 2005Keiji Hirose Abstract In order to apply the excellent chiral recognition ability of chiral pseudo-18-crown-6 ethers that we developed to chiral separation, we prepared a chiral stationary phase (CSP) by immobilizing a chiral pseudo-18-crown-6-type host on 3-aminopropyl silica gel. A chiral column was prepared by the slurry-packing method in a stainless steel HPLC column. A liquid chromatography system using this CSP combined with the detection by mass spectrometry was used for enantiomer separation of amino compounds. A normal mobile phase can be used on this CSP as opposed to conventional dynamic coating-type CSPs. Enantiomers of 18 common natural amino acids were efficiently separated. The chiral separation observed for amino acid methyl esters, amino alcohols, and lipophilic amines was fair using this HPLC system. In view of the correlation between the enantiomer selectivity observed in chromatography and the complexion in solution, the chiral recognition in host,guest interactions might contribute to this enantiomer separation. Chirality 17:142,148, 2005. © 2005 Wiley-Liss, Inc. [source] Liquid Chromatographic Separation of Olefin Oligomers and its Relation to Separation of Polyolefins , an OverviewMACROMOLECULAR SYMPOSIA, Issue 1 2009Tibor Macko Abstract Summary: Linear and branched alkanes are oligomers of polyethylene. Alkanes with higher molar masses are called waxes. These substances are widely used as fuels, oils, lubricants, etc. and for these reasons many groups have tried to analyse, separate and characterise alkanes by various methods, including liquid chromatography. Alkanes may be separated according to their size in solution by SEC. In addition to chromatographic systems separating in the SEC mode, various sorbent-solvent systems have been published, where alkanes have been separated one from another by adsorption and/or precipitation mechanism. The mobile phase is either a non-polar solvent or a polar solvent or a mixture of a solvent and a non-solvent for alkanes. Even near critical conditions, which have several advantages for applications of HPLC in polymer analysis, have been identified for alkanes. Moreover, selective separations of branched alkanes according to their structure have been published. In the majority of these published studies, solvents with low boiling points have been used as the mobile phases, which do not allow dissolution of crystalline polyolefins at atmospheric pressure. However, taking into account experiences with the separation of alkanes, new HPLC systems for the separation of polyolefins may be developed. This is a major challenge and first results are presented in this contribution. [source] | |||||||||||||