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Mobile Phase Consisting (mobile + phase_consisting)
Kinds of Mobile Phase Consisting Selected AbstractsMethodology Optimization for Quantification of Total Phenolics and Individual Phenolic Acids in Sweetpotato (Ipomoea batatas L.) RootsJOURNAL OF FOOD SCIENCE, Issue 7 2007M.S. Padda ABSTRACT:, Phenolic acids are one of the several classes of naturally occurring antioxidant compounds found in sweetpotatoes. Simplified, robust, and rapid methodologies were optimized to quantify total and individual phenolic acids in sweetpotato roots. Total phenolic acid content was quantified spectrophotometrically using both Folin,Denis and Folin,Ciocalteu reagents. The Folin,Ciocalteu reagent gave an overestimation of total phenolic acids due to the absorbance of interfering compounds (that is, reducing sugars and ascorbic acid). Individual phenolic acids were quantified by high-performance liquid chromatography (HPLC) using the latest in column technology. Four reversed-phase C18 analytical columns with different properties (dimensions, particle size, particle shape, pore size, and carbon load) were compared. Three different mobile phases using isocratic conditions were also evaluated. A column (4.6 × 150 mm) packed with 5-,m spherical silica particles of pore size 110 Å combined with 14% carbon load provided the best and fast separation of individual phenolic acids (that is, chlorogenic acid, caffeic acid, and 3 isomers of dicaffeoylquinic acid) with a total analysis time of less than 7 min. Among the 3 mobile phases tested, a mobile phase consisting of 1% (v/v) formic acid aqueous solution: acetonitrile: 2-propanol, pH 2.5 (70:22:8, v/v/v) gave adequate separation. Among the solvents tested, aqueous mixtures (80:20, solvent:water) of methanol and ethanol provided higher phenolic acid extraction efficiency than the aqueous mixture of acetone. [source] A validated liquid chromatographic/tandem mass spectrometric method for the determination of phencyclidine in microliter samples of rat serumJOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 1 2005Howard P. Hendrickson Abstract A liquid chromatographic/tandem mass spectrometric method is described for the determination of phencyclidine (PCP) in small volumes of rat serum (e.g. 50 µl). Samples were extracted using a mixed-mode strong cation-exchange column and then separated isocratically using a narrow-bore (2.1 mm i.d.) 3 µm Hypersil phenyl column and a mobile phase consisting of an ammonium formate buffer (pH 2.7) with 60% (v/v) methanol. Detection was accomplished using positive ion electrospray ionization in the multiple reaction monitoring mode. Mass spectra were obtained and peaks were observed at an m/z (% abundance) of 244 (100), 159 (25), and 86 (89). Tandem mass spectra were also obtained from the m/z 244 precursor ion with peaks observed at m/z 159 (100), 86 (96), and 91 (11). Optimum serum PCP sensitivity and precision were obtained at a transition of m/z 244 , 159. Matrix-associated ion suppression did not significantly affect the accuracy (100,112%) or precision (CV ,8%) of the assay. The lower limit of quantitation was 1 ng ml,1 in 50 µl of serum. The method was used to study the serum pharmacokinetics of PCP in rats after an intravenous bolus dose of PCP. Copyright © 2004 John Wiley & Sons, Ltd. [source] High-performance liquid chromatography determination of hydrastine and berberine in dietary supplements containing goldensealJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 7 2001Ehab A. Abourashed Abstract Goldenseal (Hydrastis canadensis L., Ranunculaceae) is an ingredient of various dietary supplements intended for enhancing general body immunity. Many goldenseal products are currently available in the United States, either alone or in combination with echinacea. In most products, the content of the main active alkaloids of goldenseal, hydrastine and berberine, is not indicated on the label. A high-performance liquid chromatography (HPLC) method has been developed for the detection and quantification of hydrastine and berberine in a number of products obtained from the United States market. The method uses a Phenomenex® Luna C18 column, a mobile phase consisting of solvent A (100 mM sodium acetate/acetic acid, pH 4.0) and solvent B (acetonitrile/methanol; 90/10, v/v). Elution was run at a flow rate of 1.0 mL/min, with a linear gradient of 80, 40% A in B over 20 min and ultraviolet detection at 290 nm. A wide range of content variation was observed for both alkaloids in the tested samples. © 2001 Wiley-Liss, Inc. and the American Pharmaceutical Association J Pharm Sci 90:817,822, 2001 [source] Simultaneous determination of yohimbine, sildenafil, vardenafil and tadalafil in dietary supplements using high-performance liquid chromatography-tandem mass spectrometryJOURNAL OF SEPARATION SCIENCE, JSS, Issue 14 2010Ying Zhang Abstract A simple and sensitive method was developed for determination of illegal adulterants (yohimbine, sildenafil, vardenafil and tadalafil) in dietary supplements by HPLC-MS/MS. The separation was achieved on a C18 column with the mobile phase consisting of acetonitrile and 0.1% acetic acid aqueous solution with a gradient elution at a flow rate of 0.5,mL/min. The analytes were quantified and identified by two characteristic transitions using the multiple-reaction monitoring mode. The recoveries of the analytes ranged from 77.5 to 109.3% with the RSD less than 8.1% (n=6). The method has been successfully applied to screen illegal adulterations of natural dietary supplements. [source] Simultaneous determination of avermectins in bovine tissues by LC-MS/MSJOURNAL OF SEPARATION SCIENCE, JSS, Issue 21 2009Koichi Inoue Abstract Analytical method for the simultaneous quantification of avermectins (AVMs), abamectin B1a, abamectin 8,9-Z isomer B1a, emamectine benzoate B1a, emamectine benzoate 8,9-Z isomer B1a, ivermectin, eprinomectin B1a, doramectin and moxidectin in bovine tissues (muscle, liver and fat) was developed by LC-MS/MS in electrospray positive ion mode. The separation was achieved on a short TSK-GEL ODS 100V column with the mobile phase consisting of acetonitrile and aquatic 0.1,mM ammonium formate containing 0.1% formic acid v/v at a flow rate of 0.2,mL/min with gradient elution. Liquid,liquid extraction with isooctane was used for the sample extraction/preparation of analytes in bovine samples. The linearity of the calibration curves was excellent in matrix-matched standards, and yielded the coefficients (r2=0.997,0.999, range from LOQ to 500, 1000 or 5000,ng/g) of determination of the target analytes. Recoveries were in the range of 87.9,99.8% with associated precision values (within-day: 1.5,7.4%, n=6, and between-day: 1.5,8.4% for 3 days) for repeatability and reproducibility. LC-MS/MS method has been proven to be highly efficient and suitable for the simultaneous determinations of eight AVMs in bovine tissue samples. [source] Determination of fluoroquinolone antibiotics in surface waters from Mondego River by high performance liquid chromatography using a monolithic columnJOURNAL OF SEPARATION SCIENCE, JSS, Issue 17 2007Angelina Pena Abstract A novel LC,fluorescence detection method based on the use of a monolithic column for the determination of norfloxacin, ciprofloxacin, and enrofloxacin antibiotic residues in environmental waters was developed. Fluoroquinolones (FQs) were isocratically eluted using a mobile phase consisting of 0.025 M phosphoric acid solution at pH 3.0 with tetrabutylammonium and methanol (960:40, v/v) through a Chromolith Performance RP-18e column (100×4.6 mm) at a flow rate of 2.5 mL/min and detected at excitation and emission wavelengths of 278 and 450 nm, respectively. After acidification and addition of EDTA, water samples were extracted using an Oasis HLB cartridge. Linearity was evaluated in the range of 0.05 to 1 ,g/mL and correlation coefficients of 0.9945 for norfloxacin, 0.9974 for ciprofloxacin, and 0.9982 for enrofloxacin were found. The limit of quantification was 25 ng/L for the three FQs. The recovery of FQs spiked into river water samples at 25, 50, and 100 ng/L fortification levels ranged from 76.5 to 91.0% for norfloxacin, 78.5 to 97.2% for ciprofloxacin, and 79.4 to 93.6% for enrofloxacin. This method was successfully applied to the analysis of water samples from the Mondego River, and ciprofloxacin and enrofloxacin residues were detected in eight water samples. [source] Identification of dimer impurities in ampicillin and amoxicillin by capillary LC and tandem mass spectrometryJOURNAL OF SEPARATION SCIENCE, JSS, Issue 3 2007Chi-Yu Lu Abstract A micro-scale liquid chromatography electrospray ionization tandem mass spectrometric method was developed for the identification of polymerized impurities in ampicillin and amoxicillin in aqueous solution. Ampicillin and amoxicillin are broad-spectrum antibiotics and widely used for the treatment of human and animal infections. In this study ampicillin, amoxicillin, and their dimers were trapped in a 5-cm capillary column containing C18 sorbents. The analytes were separated on a reversed-phase column and introduced into the mass spectrometer via a nanospray ion source. An isocratic mobile phase consisting of 1% formic acid-acetonitrile (50 : 50, v/v) was used. For identification, the fragment ions of the analytes were monitored. The aim of the present study was to develop an optimized quality control method for the analysis of high molecular weight impurities of ampicillin and amoxicillin. [source] Analysis of flunarizine in the presence of some of its degradation products using micellar liquid chromatography (MLC) or microemulsion liquid chromatography (MELC) , Application to dosage formsJOURNAL OF SEPARATION SCIENCE, JSS, Issue 2 2005Dina T. El-Sherbiny Abstract The separation of flunarizine hydrochloride (FLZ) and five of its degradation products , 1-[bis(4-fluorophenyl)methyl]-4-(3-phenyl-2-propenyl)piperazine, 4-oxide (A), bis(4-fluorophenyl)methanone (B), bis(4-fluorophenyl)methanol (C), 1-(3-phenyl-2-propenyl)piperazine (D), and 1-[bis-4-fluorophenyl) methyl] piperazine (E) , could be accomplished by reversed phase liquid chromatography using either micellar or microemulsion mobile phases. Cyanopropyl-bonded stationary phase has been used with UV detection at 254 nm. Microemulsion mobile phase consisting of 0.15 M SDS, 10% n -propanol, 1% n -octanol, and 0.3% triethylamine in 0.02 M phosphoric acid of pH 7.0, has been used for the separation of FLZ and its degradation products (B, C, D, and E). Micellar mobile phases consisting of 0.15 M sodium dodecyl sulphate (SDS), 10% n -propanol, 0.3% triethylamine (TEA) in 0.02 M phosphoric acid of pH values either 4.0 or 6.8 have been used for the separation of FLZ from its degradation products, i.e. either from (B, C, D, and E) or from (A, B, C, and D), respectively. Micellar liquid chromatography (MLC) was applied to the determination of FLZ in pure form as well as in dosage forms; the calibration graph was linear over the concentration range of 0.15,50 ,g/mL with detection limit of 0.02 ,g/mL (4.19×10,8M). [source] Identification of isomeric tropane alkaloids from Schizanthus grahamii by HPLC-NMR with loop storage and HPLC-UV-MS/SPE-NMR using a cryogenic flow probePHYTOCHEMICAL ANALYSIS, Issue 2 2006Stefan Bieri Abstract Two fully automated HPLC-NMR methods are reported and compared for the structure elucidation of four isomeric tropane alkaloids from the stem-bark of an endemic Chilean plant, Schizanthus grahamii Gill. (Solanaceae). The first approach interfaced a conventional HPLC column to NMR by means of a loop storage unit. After elution with a mobile phase consisting of deuterated water and standard protonated organic solvents, the separated analytes were momentarily stored in a loop cassette and then transferred one-at-a-time to the NMR flow probe for measurements. The second strategy combined HPLC with parallel ion-trap MS detection and NMR spectroscopy using an integrated solid-phase extraction (SPE) unit for post-column analyte trapping. The SPE cartridges were dried under a gentle stream of nitrogen and analytes were sequentially eluted and directed to a cryogenically cooled flow-probe with an NMR-friendly solvent. The structures of the four isomeric alkaloids, 3, -senecioyloxy-7, -hydroxytropane, 3, -hydroxy-7, -angeloyloxytropane, 3, -hydroxy-7, -tigloyloxytropane and 3, -hydroxy-7, -senecioyloxytropane, were unambiguously determined by combining NMR assignments with MS data. Copyright © 2005 John Wiley & Sons, Ltd. [source] Rapid determination of three anticoagulant rodenticides in whole blood by liquid chromatography coupled with electrospray ionization mass spectrometryRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 18 2006Mi-cong Jin A rapid, sensitive and selective method for the simultaneous determination of bromadiolone, flocoumafen and brodifacoum in whole blood using warfarin as internal standard (IS) by high-performance liquid chromatography coupled with electrospray ionization mass spectrometry (HPLC/ESI-MS) has been developed and validated. The target compounds were extracted from the whole blood with ethyl acetate and separated on an XDB C18 column (150,mm,×,2.1,mm i.d.,×,5,µm) by using a mobile phase consisting of 0.2% acetic acid/methanol (12/88, v/v) at a constant flow rate of 0.50 mL/min. The analytes were detected using negative ESI-MS in the selected ion monitoring (SIM) mode. The molecular ions [MH], of m/z 527, 541,523 and 307 were selected for the quantification for bromadiolone, flocoumafen, brodifacoum and the IS, respectively. The calibration curves were linear (r2,>,0.995) in the concentration range of 0.50,100.00 ng/mL. The method showed a satisfactory sensitivity (0.05,0.5 ng/mL using 200 µL blood), precision (RSD,<,11.9%), accuracy (recovery: 82.0,96.1%) and selectivity. This method was successfully applied to the determination of the analytes for the diagnoses of poisoned human beings and animals. Copyright © 2006 John Wiley & Sons, Ltd. [source] Simultaneous quantification of CTN986 and its deglycosylation products in rat serum using liquid chromatography/tandem mass spectrometryRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 11 2006Jifen Guo A quantitative method for the simultaneous determination of CTN986, a flavonol triglycoside, and its two deglycosylation products rutin and hirsutin in rat serum was developed and validated for the investigation of the pharmacokinetics of CTN986. Analytes were isolated from the serum samples (200,µL) prior to analysis by liquid chromatography/tandem mass spectrometry (LC/MS/MS) using C18 solid-phase extraction, and were separated on a Zorbax C8 reversed-phase column with an isocratic mobile phase consisting of methanol/isopropanol/water/formic acid (20:10:70:0.1, v/v/v/v). The protonated analytes generated in the positive ion mode were monitored through multiple reaction monitoring in an eletrospray ionization source. Calibration was performed by internal standardization with CTN987, a flavonoid structurally similar to CTN986, and regression curves were constructed ranging from 2 to 1000,ng/mL in 200,µL serum samples. The intra- and inter-day precision values were below 11% and accuracy was between ,2.37 and 1.4% for all quality control samples. This quantitation method was successfully applied to pharmacokinetic studies of CTN986 in rats following oral and intravenous administration. Rutin and hirsutin were not detected in rat serum. Copyright © 2006 John Wiley & Sons, Ltd. [source] Simultaneous determination of paracetamol and dextropropoxyphene in human plasma by liquid chromatography/tandem mass spectrometry: application to clinical bioequivalence studiesRAPID COMMUNICATIONS IN MASS SPECTROMETRY, Issue 6 2005Ophelia Q. P. Yin A liquid chromatography/mass spectrometry method for simultaneous determination of paracetamol and dextropropoxyphene in human plasma is described. Paracetamol and dextropropoxyphene, together with their internal standards (tolbutamide and pyrroliphene), were extracted from 0.5,mL of plasma using solid-phase extraction. The chromatography was performed using a Thermo Hypersil APS-2 Amino column (250,mm,×,4.6,mm, 5,,m) with a mobile phase consisting of acetonitrile and 0.4% glacial acetic acid in water (20:80). The total run time was 6,min for each sample. The triple-quadrupole mass spectrometer was operated in both positive (for detection of dextropropoxyphene and its IS pyrroliphene) and negative (for detection of paracetamol and its IS tolbutamide) modes using a polarity-switching technique. Multiple reaction monitoring was used for quantification. The method was linear over the concentration range of 0.1,20,,g/mL for paracetamol and 0.5,80,ng/mL for dextropropoxyphene. The intra- and inter-day precision were less than 10%, and the accuracy ranged from 92.2,110.9%. The lower limits of quantification were 0.1,,g/mL for paracetamol and 0.5,ng/mL for dextropropoxyphene. The present method provides a robust, fast and sensitive analytical tool for both paracetamol and dextropropoxyphene, and has been successfully applied to a clinical bioequivalence study in 14 subjects. Copyright © 2005 John Wiley & Sons, Ltd. [source] Sensitive and selective liquid chromatography,tandem mass spectrometry method for the determination of metoclopramide in human plasma: application to a bioequivalence studyBIOMEDICAL CHROMATOGRAPHY, Issue 9 2010Jaswanth Kumar Inamadugu Abstract A simple, sensitive and rapid method has been developed and validated for determination of the metoclopramide (MCP) in 100,,L human plasma. The analytical procedure involves a liquid,liquid extraction method using tramadol as an internal standard (IS). Chromatographic separation was carried out on a HyPURITY ADVANCE column using a mobile phase consisting of acetonitrile and 10,mm ammonium acetate buffer in the ratio of 80:20 (v/v) at a flow rate of 0.3,mL/min. The total run time of analysis was 2.5,min and elution of MCP and IS occurred at 0.9 and 1.3,min, respectively. A linear response function was established for the range of concentrations 0.53,42.07,ng/mL (r > 0.99). The intra- and inter-day precision values for MCP met the acceptance as per FDA guidelines. MCP was stable in a battery of stability studies viz., bench-top, auto-sampler and freeze,thaw cycles. The developed assay method was successfully applied to an oral bioequivalence study in humans. Copyright © 2010 John Wiley & Sons, Ltd. [source] Liquid chromatographic,tandem mass spectrometry assay for quantitation of a novel antidiabetic S002-853 in rat plasma and its application to pharmacokinetic study,BIOMEDICAL CHROMATOGRAPHY, Issue 7 2010N. Gautam Abstract A sensitive and selective LC-MS/MS method has been developed and validated for the estimation of novel antidiabetic synthetic flavonoid S002-853 in rat plasma using centchroman as an internal standard. The method involves a simple two-step liquid,liquid extraction with diethyl ether. The analyte was chromatographed on a Pierce Spheri-5, guard cyano column (30 × 4.6,,mm i.d., 5,,µm) with isocratic mobile phase consisting of methanol,ammonium acetate buffer (pH 4.6, 10,,mm; 90,:,10, v/v) at a flow rate of 0.75,,mL/min. The API 4000 triple-quadrupole LC,MS/MS system was operated under multiple reaction-monitoring mode. The ionization was performed by electrospray ionization technique in positive ion mode. The chromatographic run time was 6,,min and the weighted (1/x2) calibration curves were linear over the range 0.78,400,,ng/mL. The limit of detection and lower limit of quantification were 0.195 and 0.78,,ng/mL, respectively. The intra- and inter-batch accuracy (%bias) and precision (%RSD) were found to be less than 8.47 and 11.6% respectively. The average absolute recoveries of S002-853 and internal standard from spiked plasma samples were >90%. S002-853 was stable for 8,,h at ambient temperature, 4 weeks at ,60°C and after three freeze,thaw cycles. The assay was successfully applied to determine the pharmacokinetic parameters in male Sprague,Dawley rats after an oral dose administration at 25,,mg/kg. Copyright © 2009 John Wiley & Sons, Ltd. [source] A sensitive liquid chromatography,electrospray ionization,mass spectrometry method for the simultaneous determination of pentoxyverine citrate and guaifenesin in human plasma,application to pharmacokinetic and bioequivalence studiesBIOMEDICAL CHROMATOGRAPHY, Issue 4 2010Jinhua Wen Abstract A sensitive and specific liquid chromatography,electrospray ionization,mass spectrometry method for the identification and quantification of pentoxyverine citrate and guaifenesin in human plasma has been developed. After extraction from plasma samples by ethyl acetate, the internal standard and analytes were separated by high-performance liquid chromatographic on a Shim-pack VP-ODS C18 column (150 × 2.0 mm) using a mobile phase consisting of A (methanol) and B (0.4% glacial acetic acid and 4 mmol/L ammonium acetate) (A:B, 43 : 57). Analysis was performed on a Shimadzu LC/MS-2010A in selected ion monitoring mode with a positive electrospray ionization interface. The method was linear in the concentration range of 1.0,640.0 ng/mL for pentoxyverine citrate and 0.025,6.4 ,g/mL for guaifenesin. The inter- and intra- precision were all within 12% and accuracy ranged from 85 to 115%. The lower limits of quantification were 1.0 ng/mL for pentoxyverine citrate and 25.0 ng/mL for guaifenesin. The extraction recovery was on average 81.95% for pentoxyverine citrate and 89.03% for guaifenesin. This is the first assay method reported for the simultaneous determination of pentoxyverine citrate and guaifenesin in plasma using one chromatographic run. Copyright © 2009 John Wiley & Sons, Ltd. [source] Quantitative determination of dipyridamole in human plasma by high-performance liquid chromatography,tandem mass spectrometry and its application to a pharmacokinetic studyBIOMEDICAL CHROMATOGRAPHY, Issue 3 2010Ting Qin Abstract Dipyridamole is a classic platelet inhibitor which has been a key medicine in clinical therapy of thrombosis and cerebrovascular disease. A rapid, selective and convenient method using high-performance liquid chromatography,tandem mass spectrometry (HPLC-MS/MS) was developed for determination of dipyridamole in human plasma. After protein precipitation of 200 ,L plasma with methanol, dipyridamole and diazepam (internal standard) were chromatographed on an UltimateÔ XB-C18 (50 × 2.1 mm i.d, 3 ,) column with the mobile phase consisting of methanol,ammonium acetate (5 mM; 80 : 20, v/v) at a flow rate of 0.25 mL/min. The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring mode via positive eletrospray ionization source (ESI+). The retention times of dipyridamole and diazepam were 1.4 and 1.2 min, respectively. The method was validated over a concentration range of 0.0180,4.50 ,g/mL (r2 , 0.99) with a lower limit of quantitation (LLOQ) of 0.0180 ,g/mL for dipyridamole. The intra- and inter-day precisions (RSD) of the assay at all three QC levels were 1.6,12.7% with an accuracy (RE) of ,4.3,1.9%, which meets the requirements of the FDA guidance. The HPLC-MS/MS method herein described was proved to be suitable for pharmacokinetic study of sustained-release dipyridamole tablet in volunteers after oral administration. Copyright © 2009 John Wiley & Sons, Ltd. [source] An analytical method for cyclosporine using liquid chromatography,mass spectrometryBIOMEDICAL CHROMATOGRAPHY, Issue 2 2010Srividya V. Kanduru Abstract A liquid chromatographic mass spectrometric (LC-MS) assay has been developed for cyclosporine A (CyA) in rat plasma using amiodarone as internal standard (IS). Rat plasma (100,µL) containing drug and IS were extracted using liquid,liquid extraction with 4,mL of 95:5 ether:methanol. After evaporation of the organic layer the residue was reconstituted with 500,µL of water. Then the aqueous layer was transferred to LC-MS sample vials. A 10,µL volume was injected. The analysis was performed on a C8 column 3.5,µm (2.1 × 50,mm) heated to 60°C with a mobile phase consisting of acetonitrile:methanol:0.2% NH4OH (60:20:20) at an isocratic flow-rate of 0.2,mL/min. The ions used for quantitation of CyA and IS were m/z 1202.8 and 645.9, with retention times of 3.35 and 4.72,min, respectively. Linear relationships (r2,>,0.99) were achieved between plasma or blood concentration and peak height ratios (drug:IS) over the concentration range 50,5000,ng/mL. The CV% and mean error were <19%. Based on validation data, the lower limit of quantification for the assay was 50,ng/mL. The reported assay method displayed high measures of linearity, sensitivity, reliability and precision, allowing its applicability in pharmacokinetic studies in rat. Copyright © 2009 John Wiley & Sons, Ltd. [source] Development and validation of UPLC-MS/MS method for simultaneous determination of gestodene and ethinyl estradiol in rat plasmaBIOMEDICAL CHROMATOGRAPHY, Issue 2 2010Zhili Xiong Abstract A selective and sensitive ultra-performance liquid chromatography method with tandem mass spectrometric detection for simultaneous determination of gestodene (GES) and ethinyl estradiol (EE) in rat plasma was developed and validated. GES, EE and the internal standard, norgestrel, were extracted with ethyl acetate, derivatized (EE only) with dansyl chloride and then back-extracted into diethyl ether-hexane (2:1, v/v). The separation was performed on an ACQUITY UPLCÔ BEH C18 column with gradient elution using mobile phase consisting of acetonitrile and water (both containing 0.1% formic acid). The detection was carried out by means of electrospray ionization (ESI) mass spectrometry in positive ion mode with multiple-reaction monitoring. Calibration curves of GES and EE were linear (r2,,,0.99) over the concentration ranges 1.59,159 and 0.196,78.4,ng/mL, respectively. The intra- and inter-day precisions were not more than 6.9 and 12.9% for GES and 10.6 and 9.0% for EE, and the accuracies were ,2.5,8.0% for GES, and ,7.2,0.19% for EE, respectively. The method herein described was superior to previous methods and was applicable to the pharmacokinetic study of GES and EE in rats. Copyright © 2009 John Wiley & Sons, Ltd. [source] Simultaneous determination of six herbal components in intestinal perfusate by high-performance liquid chromatographyBIOMEDICAL CHROMATOGRAPHY, Issue 8 2009Zhanguo Wang Abstract An effective, accurate and reliable HPLC with UV detection method was developed and validated for quantitation of six components: baicalin, berberine hydrochloride, quercetin, kaempferol, isorhamnetin and baicalein in intestinal perfusate using rotundin as an internal standard. The chromatographic separation was performed on a Welchrom-C18 column (250 × 4.6 mm i.d. with 5.0 µm particle size) with a mobile phase consisting of acetonitrile, water, phosphoric acid and triethylamine (30:70:0.2:0.1,v/v) at a flow rate of 1.0 mL/min and a UV detection at 270 nm. The method had a chromatographic run time of 30 min and excellent linear behavior over the investigated concentration ranges observed with the values of r higher than 0.99 for all the analytes. The lower limit of quantification of the analytical method was 0.09 µg/mL for berberine hydrochloride, quercetin, kaempferol and baicalein and 0.18 µg/mL for baicalin and isorhamnetin. The intra- and inter-day precisions measured at three concentration levels were all less than 10% for all analytes. The bias ranged from ,6.91 to 4.33%. The validated method has been successfully applied to investigate the rat intestine absorption profiles of baicalin, berberine hydrochloride, quercetin, kaempferol, isorhamnetin and baicalein. Copyright © 2009 John Wiley & Sons, Ltd. [source] Development of a novel HPLC-MS/MS method for the determination of aconitine and its application to in vitro and rat microdialysis samplesBIOMEDICAL CHROMATOGRAPHY, Issue 7 2009Quan-long Zhang Abstract A sensitive and selective LC-MS/MS method was developed and validated for the determination of aconitine in microdialysate and rat plasma. Extraction of plasma sample was conducted by use of 1% trichloracetic acid and acetonitrile solution with 10 ng/mL internal standard (propafenone) spiked. Microdialysates were analyzed without sample purification. After sample preparation, 2 µL were injected and separated with an isocratic mobile phase consisting of acetonitrile:0.1% formic acid (60:40, v/v) at a flow rate of 0.3 mL/min. The Agilent G6410A triple quadrupole LC/MS system was operated under the multiple-reaction monitoring mode (MRM) using the electrospray ionization technique in positive mode. Overall, the assay exhibited good precision and accuracy. The diffusion properties of aconitine investigated in in vitro microdialysis experiments revealed unfavourable concentration dependence avertable by keeping a constant pH 5.77 using isotonic phosphate buffer solution as perfusate. The mean relative recoveries were 48.23% [coefficient of variation (CV 4.47%)] and 55.38% (CV 2.89%) for retrodialysis and recovery experiments, respectively. The in vivo recovery of aconitine was 34.48% (CV 3.05%) and was stable over the 6 h study period. Following characterization of aconitine both in vitro and in vivo microdialysis, the developed setting is suitable for application in pharmacokinetics and pharmacodynamics studies. Copyright © 2009 John Wiley & Sons, Ltd. [source] Development and validation of a highly sensitive and robust LC-ESI-MS/MS method for simultaneous quantitation of simvastatin acid, amlodipine and valsartan in human plasma: application to a clinical pharmacokinetic study,BIOMEDICAL CHROMATOGRAPHY, Issue 6 2009Addepalli V. Ramani Abstract A high-throughput, simple, highly sensitive and specific LC-MS/MS method has been developed for simultaneous estimation of simvastatin acid (SA), amlodipine (AD) and valsartan (VS) with 500 µL of human plasma using deuterated simvastatin acid as an internal standard (IS). The API-4000 LC-MS/MS was operated under the multiple reaction-monitoring mode (MRM) using electrospray ionization. The assay procedure involved precipitation of SA, AD, VS and IS from plasma with acetonitrile. The total run time was 2.8 min and the elution of SA, AD, VS and IS occurred at 1.81, 1.12, 1.14 and 1.81 min, respectively; this was achieved with a mobile phase consisting of 0.02 m ammonium formate (pH 4.5):acetonitrile (20:80, v/v) at a flow rate of 0.50 mL/min on an X-Terra C18 column. A linear response function was established for the range of concentrations 0.5,50 ng/mL (r > 0.994) for VS and 0.2,50 ng/mL (r > 0.996) for SA and AD. The method validation parameters for all three analytes met the acceptance as per FDA guidelines. This novel method has been applied to human pharmacokinetic study. Copyright © 2009 John Wiley & Sons, Ltd. [source] An isocratic fluorescence HPLC assay for the monitoring of l -asparaginase activity and l -asparagine depletion in children receiving E. colil -asparaginase for the treatment of acute lymphoblastic leukaemiaBIOMEDICAL CHROMATOGRAPHY, Issue 2 2009Christa E. Nath Abstract A novel assay for the determination of l -asparaginase activity in human plasma is described that is based on the HPLC quantitation of l -aspartic acid produced during enzyme incubation. Methods for monitoring l -asparagine depletion are also described. Chromatography of l -aspartic acid, l -asparagine and l -homoserine (the internal standard) involved derivatization with o -pthaldialdehyde, then separation from other amino acids on a Phenomenex Luna C18 column using a 1 mL/min flow rate and a mobile phase consisting of di-potassium hydrogen orthophosphate propionate buffer, pH 6, with 10% methanol and 10% acetonitrile. Fluoresence detection was at excitation/emission wavelengths of 357/455 nm. Under these conditions l -aspartic acid, l -asparagine and l -homoserine had retention times of 3.5, 9.8 and 17.7 min, respectively. The l -asparaginase assay was linear from 0.1 to 10 U/mL activity and interday precision and accuracy were less than 13%. The limit of quantitation was approximately 0.03 U/mL. The assay utility was established in 12 children who received E. colil -asparaginase as treatment for acute lymphoblastic leukaemia. Copyright © 2008 John Wiley & Sons, Ltd. [source] Determination of serotonin, melatonin and metabolites in gastrointestinal tissue using high-performance liquid chromatography with electrochemical detectionBIOMEDICAL CHROMATOGRAPHY, Issue 2 2009Rosanna M. W. Chau Abstract In this paper we show a simple isocratic chromatographic method for the detection of serotonin and its precursors and metabolites from various types of gastrointestinal tissue. The paper measures for the first time basal measurements of melatonin in the gastrointestinal tract, which has recently been shown to be released from the musosal lining of the gut. Tissue samples were stable following sample preparation in either 0.1 m perchloric acid or mobile phase. Analysis was carried out using a mobile phase consisting of 10% acetonitrile,90% acetate acid buffer pH 4.0 with 2 mm decane,sulfonic acid sodium salt at a column temperature of 50°C. Electrochemical detection was utilized at a potential of +850 mV vs Ag/AgCl reference electrode at 10 µA full-scale deflection. The detection limit of 5-HT and melatonin was 241 and 308 nm respectively for a 10 µL injection. As a result of the method optimization, total analysis was reduced to 30 min. Accurate responses of the tissue samples following sample preparation could be obtained following a week after storage at ,80°C. This method is capable of preparing and analysing of samples from all regions of the gastrointestinal tract. Copyright © 2008 John Wiley & Sons, Ltd. [source] Simultaneous determination of udenafil and its active metabolite, DA-8164, in human plasma and urine using ultra-performance liquid chromatography,tandem mass spectrometry: application to a pharmacokinetic studyBIOMEDICAL CHROMATOGRAPHY, Issue 9 2008Soo Kyung Bae Abstract A rapid, sensitive, and simple ultra-performance liquid chromatography,tandem mass spectrometry (UPLC/MS/MS) method for the determination of udenafil and its active metabolite, DA-8164, in human plasma and urine using sildenafil as an internal standard (IS) was developed and validated. Udenafil, DA-8164 and IS from a 100 µL aliquot of biological samples were extracted by protein precipitation using acetonitrile. Chromatographic separation was carried on an Acquity UPLC BEH C18 column (50 × 2.1 mm, i.d., 1.7 µm) with an isocratic mobile phase consisting of acetonitrile and containing 0.1% formic acid (75:25, v/v) at flow rate of 0.4 mL/min, and total run time was within 1 min. Detection and quantification was performed by the mass spectrometer using multiple reaction-monitoring mode at m/z 517 , 283 for udenafil, m/z 406 , 364 for DA-8164 and m/z 475 , 100 for IS. The assay was linear over a concentration range of 1,600 ng/mL with a lower limit of quantification of 1 ng/mL in both human plasma and urine. The coefficient of variation of this assay precision was less than 13.7%, and the accuracy exceeded 92.0%. This method was successfully applied for pharmacokinetic study after oral administration of udenafil 100 mg to healthy Korean male volunteers. Copyright © 2008 John Wiley & Sons, Ltd. [source] Optimization and validation of RP-HPLC-UV method with solid-phase extraction for determination of buparvaquone in human and rabbit plasma: application to pharmacokinetic studyBIOMEDICAL CHROMATOGRAPHY, Issue 5 2008Gantala Venkatesh Abstract A simple, sensitive and specific reversed-phase high-performance liquid chromatographic method with UV detection at 251 nm was developed for quantitation of buparvaquone (BPQ) in human and rabbit plasma. The method utilizes 250 µL of plasma and sample preparation involves protein precipitation followed by solid-phase extraction. The method was validated on a C18 column with mobile phase consisting of ammonium acetate buffer (0.02 m, pH 3.0) and acetonitrile in the ratio of 18:82 (v/v) at a flow rate of 1.1 mL/min. The calibration curves were linear (correlation coefficient ,0.998) in the selected range. The method is specific and sensitive with limit of quantitation of 50 ng/mL for BPQ. The validated method was found to be accurate and precise in the working calibration range. Stability studies were carried out at different storage conditions and BPQ was found to be stable. Partial validation studies were carried out using rabbit plasma and intra- and inter-day precision and accuracy were within 7%. This method is simple, reliable and can be routinely used for preclinical pharmacokinetic studies for BPQ. Copyright © 2007 John Wiley & Sons, Ltd. [source] Determination of huperzine A in human plasma by liquid chromatography,electrospray tandem mass spectrometry: application to a bioequivalence study on Chinese volunteersBIOMEDICAL CHROMATOGRAPHY, Issue 4 2008Wei Li Abstract A simple, sensitive and selective LC-MS-MS method has been developed for the quantification of huperzine A in human plasma. Huperzine A and pseudoephedrine hydrochloride (internal standard) were isolated from human plasma by extraction with ethyl acetate, chromatographed on a C18 column with a mobile phase consisting of 0.2% formic acid,methanol (15:85, v/v) and detected using a tandem mass spectrometer with an electrospray ionization interface. The lower limit of quantification was 0.0508 ng/mL, and the assay exhibited a linear range of 0.0508,5.08 ng/mL (r = 0.9998). The method was successfully applied to investigate the bioequivalence between two kinds of tablets (test vs reference product) in 18 healthy male Chinese volunteers. After a single 0.2 mg dose for the test and reference product, the resulting means of major pharmacokinetic parameters such as AUC0,24, AUC0,,, Cmax, Tmax and t1/2 of huperzine A were 16.35 ± 3.42 vs 16.38 ± 3.61 ng h/mL, 17.53 ± 3.80 vs 17.70 ± 3.97 ng h/mL, 2.47 ± 0.49 vs 2.51 ± 0.51 ng/mL, 1.3 ± 0.4 vs 1.2 ± 0.3 h and 5.92 ± 0.75 vs 6.18 ± 0.66 h, respectively, indicating that these two kinds of tablets were bioequivalent. Copyright © 2007 John Wiley & Sons, Ltd. [source] Simple determination of huperzine A in human plasma by liquid chromatographic,tandem mass spectrometric methodBIOMEDICAL CHROMATOGRAPHY, Issue 1 2007Yun-Xia Li Abstract Huperzine A is a potent, reversible acetylcholinesterase inhibitor. In the present work, a rapid and sensitive LC,MS,MS method for the determination of huperzine A in human plasma using codeine phosphate as internal standard has been developed and validated. The analyte and internal standard were extracted from plasma using ethyl acetate, chromatographed on a C18 column (5 µm, 150 × 4.6 mm i.d.) with a mobile phase consisting of 1% formic acid,methanol (40:60, v/v), and detected using a tandem mass spectrometer with a TurboIonSpray ionization interface. The run time was only 2 min. Good linearity was achieved in the range 0.126 -25.2 ng/mL and the limit of detection in plasma was 0.064 ng/mL. The average recovery for huperzine A was 83.4% from plasma. The analytical sensitivity and accuracy of this assay is adequate for characterization of huperzine A in human plasma. Copyright © 2006 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] Enantiomeric separation of diuretics on a novel pirkle-type chiral stationary phaseBIOMEDICAL CHROMATOGRAPHY, Issue 7 2002Ann Vercauteren A newly developed Pirkle-type straight chiral stationary phase (CSP), based on the 3,5-dinitrobenzoyl derivative of 1,2 diphenylethylene-diamine and known as ULMO, has been successfully applied to the direct resolution of the enantiomers of several diuretics by liquid chromatography. In this study, the effect of changes in the mobile phase and the intrinsic stereoselective properties of this CSP towards a specific racemate are determined experimentally. A mobile phase consisting of n-hexane and 2-propanol appears most appropriate for the chiral separation of the tested diuretics on the ULMO-column. Copyright © 2002 John Wiley & Sons, Ltd. [source] Determination of lamivudine/stavudine/efavirenz in human serum using liquid chromatography/electrospray tandem mass spectrometry with ionization polarity switchBIOMEDICAL CHROMATOGRAPHY, Issue 6 2002Bin Fan A high-performance liquid chromatography/tandem mass spectrometry (LC-MS-MS) method with ionization polarity switch was developed and validated in human serum for the determination of a lamivudine (3TC)/stavudine (d4T)/efavirenz combination HIV therapy. Solid phase extraction (SPE) was used to extract these anti-HIV drugs and internal standard aprobarbital. A gradient mobile phase consisting of acetonitrile and 20,mM ammonium acetate buffer with pH adjusted to 4.5 using glacial acetic acid was utilized to separate these drugs on a hexylsilane column (150,×,2.0,mm i.d.). The total run time between injections was 18,min. The precursor and major product ions of these drugs were monitored on a triple quadrupole mass spectrometer in the multiple reactions monitoring (MRM) mode. Ionization polarity was switched in the middle of the LC run allowing these anti-HIV drugs with different physicochemical properties to be detected simultaneously. The effect of ion suppression from human serum was studied and no interference with the analysis was noted. The method was validated over the range of 1.1,540,ng/mL for 3TC, 12.5,6228,ng/mL for d4T and 1.0,519,ng/mL for efavirenz. The method was shown to be accurate, with intra-day and inter-day accuracy less than 14.0% and precise, with intra-day and inter-day precision less than 13.1%. The extraction recoveries of all analytes were higher than 90%. Copyright © 2002 John Wiley & Sons, Ltd. [source] | ||||||||||