Separation and detection of the isomeric equine conjugated estrogens, equilin sulfate and Δ8,9-dehydroestrone sulfate, by liquid chromatography–electrospray-mass spectrometry using carbon-coated zirconia and porous graphitic carbon stationary phases

Reepmeyer, John C.; Brower, James; Ye, Hongping

doi:10.1016/j.chroma.2005.05.092

Cited by 34 publications

(22 citation statements)

References 35 publications

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“…The ability of this graphitic material to retain small polar molecules is superior to that of silica-based sorbents (Gamelin et al, 1998;Fleming et al, 1993;Remaud et al, 2005;Ayrton et al, 1995;Gu and Lim, 1990;Cantu et al, 2001;Mazan et al, 2002;Gaudin et al, 2002;Jackson and Carr, 2002;Hanai, 2003;Antonio et al, 2007;Pereira et al, 2007;Merelli et al, 2007;Tachon et al, 2007;Xia et al, 2006;Roy et al, 2006;Bohnstedt et al, 2005;Reepmeyer et al, 2005;Xing et al, 2004;Thiebaut et al, 2006;Vial et al, 2001;Lepont et al, 2001;Holmgren et al, 2005). Moreover, porous graphitic carbon proved to be chemically and pH stable, and an inert stationary phase.…”

Section: Development Of a Pgc-hplc-ms/ms Methodsmentioning

confidence: 94%

A new, validated HPLC‐MS/MS method for the simultaneous determination of the anti‐cancer agent capecitabine and its metabolites: 5′‐deoxy‐5‐fluorocytidine, 5′‐deoxy‐5‐fluorouridine, 5‐fluorouracil and 5‐fluorodihydrouracil, in human plasma

Vainchtein

Rosing

Schellens

et al. 2009

Biomedical Chromatography

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A rapid and selective liquid chromatography/tandem mass spectrometric method was developed for the simultaneous determination of capecitabine and its metabolites 5'-deoxy-5-fluorocytidine (5'-DFCR), 5'-deoxy-5-fluorouracil (5'-DFUR), 5-fluorouracil (5-FU) and dihydro-5-fluorouracil (FUH(2)) in human plasma. A 200 microL human plasma aliquot was spiked with a mixture of internal standards fludarabine and 5-chlorouracil. A single-step protein precipitation method was employed using 10% (v/v) trichloroacetic acid in water to separate analytes from bio-matrices. Volumes of 20 microL of the supernatant were directly injected onto the HPLC system. Separation was achieved on a 30 x 2.1 mm Hypercarb (porous graphitic carbon) column using a gradient by mixing 10 mm ammonium acetate and acetonitrile-2-propanol-tetrahydrofuran (1 : 3 : 2.25, v/v/v). The detection was performed using a Finnigan TSQ Quantum Ultra equipped with the electrospray ion source operated in positive and negative mode. The assay quantifies a range from 10 to 1000 ng/mL for capecitabine, from 10 to 5000 ng/mL for 5'-DFCR and 5'-DFUR, and from 50 to 5000 ng/mL for 5-FU and FUH(2) using a plasma sample of 200 microL. Correlation coefficients (r(2)) of the calibration curves in human plasma were better than 0.99 for all compounds. At all concentration levels, deviations of measured concentrations from nominal concentration were between -4.41 and 3.65% with CV values less than 12.0% for capecitabine, between -7.00 and 6.59% with CV values less than 13.0 for 5'-DFUR, between -3.25 and 4.11% with CV values less than 9.34% for 5'-DFCR, between -5.54 and 5.91% with CV values less than 9.69% for 5-FU and between -4.26 and 6.86% with CV values less than 14.9% for FUH(2). The described method was successfully applied for the evaluation of the pharmacokinetic profile of capecitabine and its metabolites in plasma of treated cancer patients.

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Section: Development Of a Pgc-hplc-ms/ms Methodsmentioning

confidence: 94%

A new, validated HPLC‐MS/MS method for the simultaneous determination of the anti‐cancer agent capecitabine and its metabolites: 5′‐deoxy‐5‐fluorocytidine, 5′‐deoxy‐5‐fluorouridine, 5‐fluorouracil and 5‐fluorodihydrouracil, in human plasma

Vainchtein

Rosing

Schellens

et al. 2009

Biomedical Chromatography

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“…Regeneration of the column was sometimes necessary to avoid loss of retention capacity, as has been reported by other groups [12, 15, 28]. The manufacturer’s instructions were used to restore the column to its original state.…”

Section: Methodsmentioning

confidence: 99%

A novel method for quantification of gemcitabine and its metabolites 2′,2′-difluorodeoxyuridine and gemcitabine triphosphate in tumour tissue by LC–MS/MS: comparison with 19F NMR spectroscopy

Bapiro

Richards

Goldgraben

et al. 2011

Cancer Chemother Pharmacol

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PurposeTo develop a sensitive analytical method to quantify gemcitabine (2′,2′-difluorodeoxycytidine, dFdC) and its metabolites 2′,2′-difluorodeoxyuridine (dFdU) and 2′,2′-difluorodeoxycytidine-5′-triphosphate (dFdCTP) simultaneously from tumour tissue.MethodsPancreatic ductal adenocarcinoma tumour tissue from genetically engineered mouse models of pancreatic cancer (KPFL/FLC and KPR172H/+C) was collected after dosing the mice with gemcitabine. 19F NMR spectroscopy and LC–MS/MS protocols were optimised to detect gemcitabine and its metabolites in homogenates of the tumour tissue.ResultsA 19F NMR protocol was developed, which was capable of distinguishing the three analytes in tumour homogenates. However, it required at least 100 mg of the tissue in question and a long acquisition time per sample, making it impractical for use in large PK/PD studies or clinical trials. The LC–MS/MS protocol was developed using porous graphitic carbon to separate the analytes, enabling simultaneous detection of all three analytes from as little as 10 mg of tissue, with a sensitivity for dFdCTP of 0.2 ng/mg tissue. Multiple pieces of tissue from single tumours were analysed, showing little intra-tumour variation in the concentrations of dFdC or dFdU (both intra- and extra-cellular). Intra-tumoural variation was observed in the concentration of dFdCTP, an intra-cellular metabolite, which may reflect regions of different cellularity within a tumour.ConclusionWe have developed a sensitive LC–MS/MS method capable of quantifying gemcitabine, dFdU and dFdCTP in pancreatic tumour tissue. The requirement for only 10 mg of tissue enables this protocol to be used to analyse multiple areas from a single tumour and to spare tissue for additional pharmacodynamic assays.

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“…The general loss of retention on PGC is thought to be mainly due to the column’s particularly high susceptibility to contamination, 7 hence the suggestion to backflush regularly. Wash procedures involve the use of strong acids/bases and solvents such as tetrahydrofuran, acetone, trifluoroacetate, hydrochloric acid, and sodium hydroxide.…”

mentioning

confidence: 99%

Understanding the Complexity of Porous Graphitic Carbon (PGC) Chromatography: Modulation of Mobile-Stationary Phase Interactions Overcomes Loss of Retention and Reduces Variability

2016

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Porous graphitic carbon (PGC) is an important tool in a chromatographer’s armory that retains polar compounds with mass spectrometry (MS)-compatible solvents. However, its applicability is severely limited by an unpredictable loss of retention, which can be attributed to contamination. The solutions offered fail to restore the original retention and our observations of retention time shifts of gemcitabine/metabolites on PGC are not consistent with contamination. The mobile phase affects the ionization state of analytes and the polarizable PGC surface that influences the strength of dispersive forces governing retention on the stationary phase. We hypothesized that failure to maintain the same PGC surface before and after running a gradient is a cause of the observed retention loss/variability on PGC. Herein, we optimize the choice of mobile phase solvent in a gradient program with three parts: a preparatory phase, which allows binding of analytes to column; an elution phase, which gives the required separation/peak shape; and a maintenance phase, to preserve the required retention capacity. Via liquid chromatography/tandem mass spectrometry (LC-MS/MS) analysis of gemcitabine and its metabolites extracted from tumor tissue, we demonstrate reproducible chromatography on three PGC columns of different ages. This approach simplifies use of the PGC to the same level as that of a C-18 column, removes the need for column regeneration, and minimizes run times, thus allowing PGC columns to be used to their full potential.

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Separation and detection of the isomeric equine conjugated estrogens, equilin sulfate and Δ8,9-dehydroestrone sulfate, by liquid chromatography–electrospray-mass spectrometry using carbon-coated zirconia and porous graphitic carbon stationary phases

Cited by 34 publications

References 35 publications

A new, validated HPLC‐MS/MS method for the simultaneous determination of the anti‐cancer agent capecitabine and its metabolites: 5′‐deoxy‐5‐fluorocytidine, 5′‐deoxy‐5‐fluorouridine, 5‐fluorouracil and 5‐fluorodihydrouracil, in human plasma

A new, validated HPLC‐MS/MS method for the simultaneous determination of the anti‐cancer agent capecitabine and its metabolites: 5′‐deoxy‐5‐fluorocytidine, 5′‐deoxy‐5‐fluorouridine, 5‐fluorouracil and 5‐fluorodihydrouracil, in human plasma

A novel method for quantification of gemcitabine and its metabolites 2′,2′-difluorodeoxyuridine and gemcitabine triphosphate in tumour tissue by LC–MS/MS: comparison with 19F NMR spectroscopy

Understanding the Complexity of Porous Graphitic Carbon (PGC) Chromatography: Modulation of Mobile-Stationary Phase Interactions Overcomes Loss of Retention and Reduces Variability

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